Plasticity in gene expression underlies much of the ability of organisms to cope with environmental variation. Understanding limits in the ability of organisms to adjust to a range of environments has implications for understanding how pathogens respond to human environments (e.g., antibiotic resistance), and how humans respond, at times inappropriately, to current environments (e.g., obesity). This research tests the hypothesis that the degree of modularity, or environmental specificity, of gene expression has fundamental implications for defining limits in the evolution of plasticity. Highly modular gene expression is assumed to permit adaptation to specific environments because overlap in gene function, and thus pleiotropy, is much reduced. At the same time, highly modular gene expression comes at a potential cost: when alternative environments are experienced infrequently, selection on environment-specific genes is weakened and mutation accumulation can occur. These hypotheses will be tested in a newly emerging model system in evolutionary developmental genetics: homed beetles in the genus Onthophagus. These insects display highly divergent alternative phenotypes depending on larval nutrition. Such alternative morphs develop through expression of genes shared across the development of traits, as well as genes specific to different morphs. This research tests several predictions by contrasting the development of secondary sexual traits (horns) and brains in horned (aggressive fighter) and hornless (non-aggressive sneaker) beetle morphs. This research tests four predictions both within and between species: 1) increased environmentally-specific gene expression will be correlated with the evolution of greater ranges of plasticity, 2) genes specific to the development of morph behavior and morphology will be more genetically variable than those shared between morphs, 3) genes specific to morph development experience higher rates of genetic divergence as a result of weak selection, and 4) morph-specific genes will play functional roles in generating divergence between morphs. This work will use a variety of tools developed for Onthophagus beetles, including transcriptional profiling via microarray analysis, and investigating gene function through RNA interference gene knock-down. Relevance. Understanding evolutionary limits of plasticity in gene expression has implications for public health. If we can curb the ability of pathogens to cope with multiple environments, such as invading the human body or dealing with multiple medications, we may be able to reduce pathogen virulence. We may learn why humans cannot express genes that promote health in a wide range of environments, such as diabetes-inducing nutritional environments or changes in the aging brain. [unreadable] [unreadable] [unreadable]